Characterization of charge‐variant bioconjugates derived from chemical labeling of proteins

Abstract

Purified antibodies and other target‐specific recognition proteins are commonly used reagents in a wide range of biochemical analyses from basic research to the clinical laboratory. Characterization of charge states for these purified protein reagents is often critical to indication of solution‐phase stability and the potential for non‐specific interactions with solid‐phase materials and other proteins. Conjugation, or chemical labeling, with various small molecules adds further complexity to the inherent charge‐variation from isoforms of a single purified protein. While these added labels typically cause minimal change in the overall size of the protein molecule and infrequently inhibit the native protein function, they can cause a significant change in effective protein charge. The experiments described here examine the effect of an amine‐reactive protein labeling process on the charge state distribution of resulting bioconjugates. Various analytical methods were applied to resolve and characterize the changes in protein charge state distribution as a function of the labeling load (label‐to‐protein ratio, or incorporation ratio (IR)) and as a function of the type of label used. A distinction between surface charge effects and net protein charge effects is addressed.